Project Abstract Progress in glioma treatment has been disappointing owing to our poor understanding of the underlying biology. A glioma contains not only cancer cells but also a variety of non-glioma brain cells including glia. However, the interaction between glia and glioma cells and the contribution of glia to glioma growth is largely unexplored. We have developed immunocompromised mice, which will make it possible to xenograft human or transplant mouse glioma cells into the brain of these mice and to trace different non-glioma cell types that have received extracellular vesicles from glioma cells. With this method, we will investigate the mechanisms underlying the crosstalk between glioma cells and glia, including adjacent astrocytes, and the role of glioma- adjacent astrocytes in glioma development. The main challenge to studying astrocyte-glioma cell interactions in vivo is to specifically label both cell types and observe them simultaneously with high-resolution microscopy. We can genetically tag astrocytes with red tdTomato for fate mapping after transplantation of human- or mouse-derived glioma cells. We have developed a longitudinal time-lapse imaging method to directly observe GFP-expressing glioma cells in the same transgenic mouse from days to weeks. These methods allow us to characterize the fate of astrocytes and demonstrate how astrocytes form a unique tumor microenvironment (TME), which is critical for glioma progression. We will further verify these results in both syngeneic mouse model and somatic engineered mouse model. This work will provide the brain tumor research community with (1) a novel mechanism of glioma-glial cell interactions and (2) a cellular therapeutic target?astrocytes adjacent to gliomas?for manipulating glioma growth.